感染“褐色沉积症”虾夷扇贝的外套膜转录组测序及差异表达基因分析

Transcriptome sequencing and analysis of differentially expressed genes in the mantle tissues of scallops(Mizuhopecten yessoensis)infected with Brown Deposition Syndrome

为分析“褐色沉积症”形成分子机制,探究筏式养殖虾夷扇贝(Mizuhopecten yessoensis)贝壳内侧褐色沉积症状形成原因,采用Illumina高通量测序平台对感染“褐色沉积症”虾夷扇贝和健康虾夷扇贝外套膜组织进行转录组测序分析。结果显示,患病和健康扇贝外套膜分别获得43862251和41806737条clean reads。经参考基因组比对分析,患病和健康扇贝外套膜表达基因数量分别为17835个和16816个,差异表达基因208个,其中上调基因170个,下调基因38个。选取6个差异表达基因进行荧光定量PCR(qRT-PCR)验证,结果证明转录组测序分析可靠。GO功能富集发现差异基因主要富集在几丁质代谢、含氨基葡萄糖化合物代谢、几丁质结合和氨基糖代谢等与几丁质合成代谢相关的通路;KEGG通路富集分析发现差异基因主要参与内质网蛋白质合成、内吞作用、剪接体和谷胱甘肽代谢等途径;对差异表达倍数显著的前40个基因分析发现,编码类咖啡酰辅酶A-O-甲基转移酶(caffeoyl-CoA O-methyltransferase-like,CCOAOMT-L)、类1-氨基环丙烷-1羧酸合酶(1-aminocyclopropane-1-carboxylate synthaselike protein,ACCS-L)、黄蛋白(protein yellow-like)、热休克蛋白70(HSP70)、过氧化物酶成熟因子(DUOXA1)、多烯脂肪酸异构酶(poly enoic fatty acid isomerase,PFI)、Na^(+)/Cl^(+)依赖的甘氨酸转运蛋白(sodium-and chloride-dependent glycine transporter,GlyT1)、类酪氨酸酶(putative tyrosinase-like protein,Putative Tyr-3)、Toll样受体(protein toll)等与蛋白质醌鞣化、免疫、氧化、神经传递相关的基因与褐色沉积症形成密切相关,揭示了虾夷扇贝外套膜褐色沉积症形成的内在原因。

During the past few years,outbreaks of brown deposition syndrome have caused massive mortality in the raft culture scallop,Mizuhopecten yessoensis,in Changhai,Liaoning.The moribund scallops exhibited severe disease signs including brown deposits on the inner shell.To analyze the molecular mechanism of brown deposit symptom and find the key genes,the transcriptomes of mantle tissues of scallop M.yessoensis infected with“Brown Deposition Syndrome”and healthy scallops were sequenced using the Illumina high-throughput sequencing platform,and the differentially expressed genes(DEGs)were analyzed.A total of 43862251 and 41806737 clean reads were obtained in the mantle tissue of diseased and healthy scallops.A total of 17835 genes and 16816 genes were obtained from the reference genome analysis.The degs were screened out with a threshold criterion of|log2(FoldChange)|>0 and P-value<0.005.A total of 208 genes were identified,among which 170 genes were up-regulated and 38 genes genes were down-regulated.Six functional DEGs were randomly selected for qRT-PCR analysis,and the results confirmed that the transcriptome analysis was reliable.The result of gene ontology(GO)functional enrichment showed that the DEGs were significantly enriched in the chitin metabolic process,the glucosamine-containing compound metabolic process,the chitin binding process,and the amino sugar metabolic process.KEGG pathway enrichment suggested that the DEGs were enriched in protein processing in the endoplasmic reticulum,endocytosis,spliceosome and glutathione metabolism.The top 40 DEGs in NR,NT and Swissprot were identified and analyzed.The genes CCOAOMT-L,ACCS-L,protein yellow-like,HSP70,DUOXA1,PFI,GlyT1,Putative tyrosinase-like protein Tyr-3,and protein Toll have been linked to Brown Deposition Syndrome.Protein Quinone tanning,ROS metabolism,lipid peroxidation,the immune system,and neurotransmission are regulated by these genes.